Methods for monitoring and guiding therapeutic suppression of parathyroid hormone in renal patients having secondary hyperparathyroidism

ABSTRACT

The present invention relates to kits for monitoring and guiding therapeutic suppression of parathyroid hormone in renal patients having secondary hyperparathyroidism. The kit provides means to measure the level of a PTH agonist and a PTH antagonist in a patient, and it further provides means for administering a PTH suppressing therapeutic to the patient so as to reduce the level of a PTH agonist and minimize the level of parathyroid hormone antagonist.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a divisional of U.S. patent application Ser. No.10/286,465, filed Nov. 1, 2002, now allowed, which is a continuation inpart of U.S. patent application Ser. No. 10/002,818, filed Nov. 2, 2001,now issued as U.S. Pat. No. 6,524,788, each of which is incorporatedherein by reference.

TECHNICAL FIELD

The present invention relates to novel methods for monitoring andguiding therapeutic suppression of parathyroid hormone in renal patientshaving secondary hyperparathyroidism. One determines and monitors thelevel of parathyroid hormone agonist and parathyroid hormone antagonistin the renal patient. The parathyroid hormone suppressing therapeutic isadministered to the patient so as to minimize the level of parathyroidhormone antagonist.

BACKGROUND ART

Calcium plays an indispensable role in cell permeability, the formationof bones and teeth, blood coagulation, transmission of nerve impulse,and normal muscle contraction. The concentration of calcium ions in theblood is, along with calcitrol and calcitonin, regulated mainly byparathyroid hormone (PTH). Although calcium intake and excretion mayvary, PTH serves through a feedback mechanism to maintain a steadyconcentration of calcium in cells and surrounding fluids. When serumcalcium lowers, the parathyroid glands secrete PTH, affecting therelease of stored calcium. When serum calcium increases, stored calciumrelease is retarded through lowered secretions of PTH.

The complete form of human PTH, sometimes referred to in the art as hPTHbut referred to in the present invention as an example of PTH agonist,is a unique 84 amino acid peptide (SEQ ID NO: 1), as is shown in FIG. 1.Researchers have found that this peptide has an anabolic effect on bonethat involves a domain for protein kinase C activation (amino acidresidues 28 to 34) as well as a domain for adenylate cyclase activation(amino acid residues 1 to 7). However, various catabolic forms ofclipped or fragmented PTH peptides also are found in circulation, mostlikely formed by intraglandular or peripheral metabolism. For example,whole PTH can be cleaved between amino acids 34 and 35 to produce a(1-34) PTH N-terminal fragment and a (35-84) PTH C-terminal fragment.Likewise, clipping can occur between either amino acids 36 and 37 or 37and 38. Recently, a large PTH fragment referred to as “non-(1-84) PTH”has been disclosed which is clipped closer to the N-terminal end of PTH.(See R. LePage et al., “A non-(1-84) circulating parathyroid hormone(PTH) fragment interferes significantly with intact PTH commercial assaymeasurements in uremic samples,” Clin. Chem. ( 1998 ); 44: 805-810.)

The clinical need for accurate measurement of PTH is well demonstrated.Serum PTH level is one of the most important indices for patients withthe following diseases: familial hypocalciuria; hypercalcemia; multipleendocrine neoplasia types I and II; osteoporosis; Paget's bone disease;primary hyperparathyroidism—caused by primary hyperplasia or adenoma ofthe parathyroid glands; pseudohypoparathyroidism; and renal failure,which can cause secondary hyperparathyroidism.

PTH plays a role in the course of disease in a patient with chronicrenal failure. Renal osteodystrophy (RO) is a complex skeletal diseasecomprising osteitis fibrosa cystica (caused by PTH excess),osteomalacia—unmineralized bone matrix (caused by vitamin D deficiency),extraskeletal calcification/ossification (caused by abnormal calcium andphosphorus metabolism), and adynamic low bone turnover disease(contributed to by PTH suppression). Chronic renal failure patients candevelop RO. Failing kidneys increase serum phosphorus hyperphosphoremia)and decrease 1,25-dihydroxyvitamin D (1,25-D) production by the kidney.The former results in secondary hyperparathyroidism from decreasedgastrointestinal calcium absorption and osteitis fibrosa cystica fromincreased PTH in response to an increase in serum phosphorus. The latercauses hypocalcemia and osteomalacia. With the onset of secondaryhyperparathyroidism, the parathyroid gland becomes less responsive toits hormonal regulators because of decreased expression of its calciumand vitamin D receptors. Serum calcium drops. RO can lead to digitalgangrene, bone pain, bone fractures, and muscle weakness.

For chronic renal failure patients with secondary hyperparathyroidism, anumber of different therapeutic treatments are available. One canadminister calcium carbonate so as to directly adjust the availablecalcium ion level. However, with the increasing incidence of ectopiccalcification, increasing calcium intake is often not desirable. One canadminister calcimimetics, such as AMG073 made by Amgen, Inc. of ThousandOaks, Calif. However, AMG073 has not been approved for use in the USA.One can administer vitamin D analogues, (such as the Calcijex® orZemplar® brands made by Abbott Labs of Abbott Park, Ill.; Rocaltrol®brand made by Roche Laboratories of Basle, Switzerland; Oxarol brandmade by Chugai Pharmaceutical), so as to lower PTH. However, researchershave found that vitamin D analogues can oversuppress PTH, therebyleading to adynamic low bone turnover disease setting the patient atrisk of ectopic and vascular calcification. (See the package insert forZemplar®, Abbott Reference 06-9998-R1-Rev, April 1998. See the packageinsert for Rocaltrol®, Roche Laboratories, inc. November 1998 Productidentification Guide, page 334.)

Researchers have also found that a large circulating PTH fragment (e.g.,cyclase inactive parathyroid hormone) functions as a naturally occurringPTH antagonist. Cyclase inactive PTH has been found to be useful,alongside whole PTH, as an indicator in separating untreated end stagerenal disease (ESRD) patients with high bone turnover from those withadynamic low bone turnover. (See Faugere, M. C. et alia. “ImprovedAssessment of Bone Turnover by the PTH 1-84/large C-PTH fragments ratioin ESRD patients”, Kidney International 2001; 60: 1460-1468.) Moreover,researchers have found that cyclase inactive PTH can cause adynamic lowbone turnover by inhibiting the formation of osteoclasts, boneresorption, and bone turnover. (See Divieti, P. et alia, “In vitroInhibition of Bone Resorption by Human PTH (7-84)” J. Bone Miner Res2001 :Suppl 1, S307. See also Faugere, M. C. et alia, “The Effects ofPTH (1-84) on bone turnover are Antagonized by PTH (7-84) inThyroparathyroidectomized and Nephrectomized Rats”; J Am Soc Nephrol12:2001, 764A.)

Determining circulating biologically active PTH levels in humans hasbeen challenging. One major problem is that PTH is found at low levels,normally 10 pg/mL to 65 pg/mL. Coupled with extremely low circulatinglevels is the problem of the heterogeneity of PTH and its manycirculating fragments. In many cases, immunoassays have facedsubstantial and significant interference from circulating PTH fragments.For example, some commercially available PTH kits have almost 100%cross-reactivity with the non- (1-84) PTH fragment, (see the LePagearticle).

PTH immunoassays have varied over the years. One early approach is adouble antibody precipitation immunoassay found in U.S. Pat. No.4,369,138 to Arnold W. Lindall et alia. A first antibody has a highaffinity for a (65-84) PTH fragment. A radioactive labeled (65-84) PTHpeptide is added to the sample with the first antibody to compete forthe endogenous unlabeled peptide. A second antibody is added which bindsto any first antibody and radioactive labeled PTH fragment complex,thereby forming a precipitate. Both precipitate and supernatant can bemeasured for radioactive activity, and endogenous PTH levels can becalculated therefrom.

In an effort to overcome PTH fragment interference, immunoradiometrictwo-site assays for intact PTH (I-PTH) have been introduced, such asAllegro® Intact PTH assay by the Nichol's Institute of San JuanCapistrano, Calif. In one version, a capture antibody specifically bindsto the C-terminal portion of hPTH while a labeled antibody specificallybinds to the N-terminal portion of the captured hPTH. In another, twomonoclonal antibodies were used, both of which attached to theN-terminal portion of hPTH. Unfortunately, these assays have problems inthat they measure but do not discriminate between whole PTH andnon-whole PTH peptide fragments. This inability comes to the fore inhyperparathyroid patients and renal failure patients who havesignificant endogenous concentrations of large, non-whole PTH fragments.

Recently, Scantibodies Laboratory, Inc. of Santee, Calif. USA introduceda series of novel kits that allow for the accurate measurement of bothPTH agonist and PTH antagonist. The PTH agonist assay is a directmeasurement, while the PTH antagonist assay is a calculated value fromthe difference of the PTH agonist assay value and a total PTH (includingboth PTH agonist and PTH antagonist) assay value. A number of unexpectedadvantages have become available to the physician, including the firstnon-invasive method for assisting in the differentiation of secondaryhyperparathyroid patients with HBT and ALBT.

DISCLOSURE OF THE INVENTION

In one embodiment, a method is provided for monitoring and guidingtherapeutic suppression of parathyroid hormone in renal patients havingsecondary hyperparathyroidism comprising: a) determining and monitoringthe level of parathyroid hormone (PTH) agonist in a renal patient havingsecondary hyperparathyroidism; b) determining and monitoring the levelof PTH antagonist in the patient; and c) administering a therapeutic tothe patient that suppresses PTH agonist whereby the amount oftherapeutic administered is adjusted such that the level of PTHantagonist is minimized. Frequently, the therapeutic is administered inincreasing increments from a nominal amount. In addition, frequentlysteps a) and b) are performed using a sample obtained from a renalpatient, which sample may be a serum, plasma and/or blood sample.

In one aspect the renal patient is already receiving the therapeutic,and therapeutic administration is terminated for a time sufficient toallow the patient to return to at least a relatively non-suppressedstate. In one aspect, this timing may be measured with respect toparathyroid gland status and/or bone status. Relatedly, the timesufficient to allow the patient to return to at least a relativelynon-suppressed state is frequently between about three months to aboutsix months, with respect to bone status. The time sufficient to allowthe patient to return to at least a relatively non-suppressed state isfrequently between about three minutes to about twenty minutes, withrespect to parathyroid gland status. On occasion, the time sufficient toallow the patient to return to at least a relatively non-suppressedstate is between about two weeks to about six weeks, or sometimes aboutfour weeks. In addition, while guiding therapy of these patients, theamount of administered therapeutic may be adjusted until the level ofPTH antagonist is minimized.

In another aspect of the present invention the therapeutic beingadministered is selected from the group consisting of calcium, vitaminD, vitamin D analogues, and calcimimetics (e.g., AMG073). Frequently,when vitamin D analogs are used, they may be selected fromparicalcitrol, calcitriol, maxacalcitol, alfacalcidol, calcifediol, orergocalciferol.

In a further aspect of the present invention, the PTH agonist level iscompared with the PTH antagonist level. In another aspect, the PTHagonist level is compared with the total parathyroid hormone level. Inyet another aspect, the PTH antagonist level is compared with the totalparathyroid hormone level. In a further aspect, the PTH agonist level iscompared with the PTH antagonist level in the form of a ratio orproportion.

In one aspect, the PTH antagonist level is determined by determining atotal PTH level and determining a PTH agonist level followed bysubtracting the PTH agonist level from the total PTH level.

In another aspect, the PTH agonist level and the PTH antagonist levelare determined using an immunoassay. In yet another aspect, the PTHagonist level is determined using an antibody that distinguishes PTHagonist from PTH antagonist. In a further aspect, frequently the PTHantagonist level is determined using an antibody that distinguishes PTHagonist from PTH antagonist.

In one embodiment, the method above may further comprise monitoring forvascular calcification in the patients. Frequently, vascularcalcification is monitored by monitoring alkaline phosphatase levels inthe patient.

In another embodiment, the PTH agonist levels and the PTH antagonistlevels are compared, individually or together, with corresponding bonebiopsy data. In one aspect, the ratio or proportion of PTH agonist toPTH antagonist will be compared with corresponding bone biopsy data. Onoccasion, bone biopsy data will be used to verify the results of PTHagonist level and/or PTH antagonist level determinations and/or PTHagonist/antagonist ratios. Frequently, the PTH agonist level and the PTHantagonist level determinations are compared with corresponding bonebiopsy data after administration of the therapeutic. On occasion, thiscomparison after administration of the therapeutic is useful to monitorthe PTH therapy of patients. Frequently, such treatment is directed todecreasing PTH agonist levels and minimizing PTH antagonist levels.

In a further aspect, the PTH agonist comprises a contiguous portion ofhuman PTH having an amino acid sequence set forth in SEQ ID NO:1(PTH₁₋₈₄), and the PTH agonist has the following characteristics: theN-terminal amino acid residue of the PTH agonist starts at position 1 ofthe PTH₁₋₈₄; and the C-terminal amino acid residue of the PTH agonistends at any position spanning position 34 through position 84 of thePTH₁₋₈₄. In another aspect, the PTH antagonist comprises a contiguousportion of human PTH having an amino acid sequence set forth in SEQ IDNO: 1 (PTH₁₋₈₄), and the PTH antagonist has the followingcharacteristics: the N-terminal amino acid residue of the PTH antagoniststarts at any position spanning position 2 through position 33 of thePTH₁₋₈₄; the C-terminal amino acid residue of the PTH antagonist ends atany position spanning position 35 through position 84 of the PTH₁₋₈₄;and the PTH antagonist has a minimal length of three amino acidresidues.

In another embodiment, a method is provided for monitoring and guidingtherapeutic suppression of parathyroid hormone in renal patients havingsecondary hyperparathyroidism comprising: a) obtaining a sample from arenal patient having secondary hyperparathyroidism; b) determining andmonitoring the level of PTH antagonist in the renal patient; and c)administering a therapeutic to the patient capable of suppressing a PTHagonist, whereby the amount of therapeutic administered is adjusted suchthat the level of PTH antagonist is minimized in the patient.

In a further embodiment, kits are provided that are useful formonitoring and guiding therapeutic suppression of parathyroid hormone inrenal patients having secondary hyperparathyroidism comprising: a) meansfor monitoring the level of parathyroid hormone (PTH) agonist in a renalpatient having secondary hyperparathyroidism; b) means for monitoringthe level of PTH antagonist in the patient; and c) means foradministering a therapeutic to the patient that suppresses PTH agonistwhereby the amount of therapeutic administered is adjusted such that PTHagonist levels are decreased and the level of PTH antagonist isminimized.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a diagrammatic view of human wPTH.

FIG. 2 is a graph comparing PTH measurement parameters over time forpatients receiving a PTH suppressant therapy.

DETAILED DESCRIPTION OF THE INVENTION

For clarity of disclosure, and not by way of limitation, the detaileddescription of the invention is divided into the subsections thatfollow.

A. Definitions

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as is commonly understood by one of ordinary skillin the art to which this invention belongs. All patents, applications,published applications and other publications referred to herein areincorporated by reference in their entirety. If a definition set forthin this section is contrary to or otherwise inconsistent with adefinition set forth in the patents, applications, publishedapplications and other publications that are herein incorporated byreference, the definition set forth in this section prevails over thedefinition that is incorporated herein by reference.

As used herein, “a” or “an” means “at least one” or “one or more.”

As used herein, “parathyroid hormone (PTH) agonist” refers to thecomplete molecule of PTH or a fragment, derivative or analog thereofthat stimulates osteoclasts formation and bone turnover to increaseblood calcium levels. PTH agonist further refers to peptides which havePTH agonist properties. Other names of PTH include parathormone andparathyrin. For purposes herein, the name “parathyroid hormone (PTH)” isused herein, although all other names are contemplated. It is intendedto encompass PTH agonist with conservative amino acid substitutions thatdo not substantially alter its biological activity. Suitableconservative substitutions of amino acids are known to those of skill inthis art and may be made generally without altering the biologicalactivity of the resulting molecule. Those of skill in this art recognizethat, in general, single amino acid substitutions in non-essentialregions of a polypeptide do not substantially alter biological activity(see, e.g., Watson et al., MOLECULAR BIOLOGY OF THE GENE, 4th Edition,1987, The Bejamin/Cummings Pub. Co., p.224). PTH agonist assay valuesmay be obtained by measuring a sample with a Scantibodies Whole PTHAssay or a Scantibodies CAP Assay or a 3^(rd) generation PTH Assay or aNichols BioIntact PTH assay or an Immutopics Human Bioactive PTH assay.“Cyclase activating PTH,” “whole PTH,” and “CAP” are representativeexamples of PTH agonists.

As used herein, “parathyroid hormone (PTH) antagonist” refers to a PTHfragment or derivative that counters the effect of a PTH agonist orotherwise lacks PTH agonist activity. It is intended to encompass PTHantagonist with conservative amino acid substitutions that do notsubstantially alter its activity. Suitable conservative substitutions ofamino acids are known to those of skill in this art and may be madegenerally without altering the biological activity of the resultingmolecule. Those of skill in this art recognize that, in general, singleamino acid substitutions in non-essential regions of a polypeptide donot substantially alter biological activity (see, e.g., Watson, et al.MOLECULAR BIOLOGY OF THE GENE, 4th Edition, 1987, The Bejamin/CummingsPub. co., p.224). “Cyclase inactive PTH” and “CIP” are representativeexamples of PTH antagonists.

As used herein, the terms “total PTH,” “intact PTH” and “total intactPTH” are interchangeable and refer to an assay directed at measuring PTHagonist and PTH antagonist levels.

As used herein, a “functional derivative or fragment” of PTH agonist orPTH antagonist refers to a derivative or fragment of PTH that stillsubstantially retains its function as a PTH agonist or PTH antagonist.Normally, the derivative or fragment retains at least 50% of its PTHagonist or PTH antagonist activity. Preferably, the derivative orfragment retains at least 60%, 70%, 80%, 90%, 95%, 99% and 100% of itsPTH agonist or PTH antagonist activity. It is also possible that afunctional derivative or fragment of PTH agonist or PTH antagonist hashigher PTH agonist or PTH antagonist activity than a parent moleculefrom which the functional derivative or fragment is derived from.

As used herein, “treatment” means any manner in which the symptoms of acondition, disorder or disease are ameliorated or otherwise beneficiallyaltered. Treatment also encompasses any pharmaceutical use of thecompositions herein.

As used herein, “disease or disorder” refers to a pathological conditionin an organism resulting from, e.g., infection or genetic defect, andcharacterized by identifiable symptoms.

As used herein, “adynamic low bone turnover disease” refers to a varietyof disorders involving abnormal PTH agonist and/or antagonist levels ina person. This definition is non-limiting in that it does not refer toonly one specific disease, it refers to a variety of disorders that mayresult from abnormal PTH or PTH component levels in a person. As PTHlevels are tied to bone turnover rate, abnormally low levels of PTHagonist, abnormally low levels of PTH agonist/antagonist ratios, andabnormally high levels of PTH antagonist may lead to abnormally low boneturnover in a person. In a person, this type of state may indicate thepresence of, or susceptibility to, an adynamic low bone turnoverdisease. Conversely, abnormally high levels of PTH agonist, abnormallyhigh levels of PTH agonist/antagonist ratios, and abnormally low levelsof PTH antagonist may lead to abnormally high bone turnover in a person.

As used herein the term “sample” refers to anything which is suspectedof containing an analyte for which an analyte assay is desired. Thesample may be a biological sample, such as a biological fluid or abiological tissue. Examples of biological fluids include urine, blood,plasma, serum, saliva, semen, stool, sputum, cerebral spinal fluid,tears, mucus, amniotic fluid or the like. Biological tissues areaggregate of cells, usually of a particular kind together with theirintercellular substance that form one of the structural materials of ahuman, animal, plant, bacterial, fungal or viral structure, includingconnective, epithelium, muscle and nerve tissues. Examples of biologicaltissues also include organs, tumors, lymph nodes, arteries andindividual cell(s). Frequently, samples obtained for use in the presentinvention contain, or are suspected of containing, levels of PTH agonistand/or PTH antagonist that are detectable through methods described andcontemplated herein.

As used herein, the term “vitamin D analog” refers to any availablesource of synthetic vitamin D from a variety of sources for clinical orexperimental use. For example, vitamin D analogs may be obtained fromsources such as Abbott Laboratories, Bone Care International, Hoffman-LaRoche, Ltd., Chugai Pharmaceutical Co., Amgen, Inc., NPSPharmaceuticals, Inc, Kirin Brewing Company, Ltd., Sumitomo Corp., etc.Examples of vitamin D analogs available from these sources includeCalcijex®, Zemplar®, Hectoral®, Rocaltrol®, Oxarol, etc. In addition,the active ingredients in a variety of synthetic vitamin D sourcescontemplated herein may include paricalcitrol, calcitriol, maxacalcitol,alfacalcidol, calcifediol, or ergocalciferol.

B. Risk and therapy determinations based on PTH agonist levels and PTHantagonist levels

The present invention relates to novel methods for monitoring andguiding therapeutic suppression of parathyroid hormone in renal patientshaving secondary hyperparathyroidism. In one aspect, one determines andmonitors the level of PTH agonist and PTH antagonist in the renalpatient. In another aspect, one determines and monitors the level of PTHagonist in the renal patient. Frequently, the parathyroid hormonesuppressing therapeutic is administered to the patient so as to minimizethe level of PTH antagonist.

Secondary hyperparathyroidism is a common disease in renal compromisedpatients, especially those with ESRD. Virtually all ESRD patients havebone disease and mineral metabolism disorders, either high bone turnoverdisease or adynamic low bone turnover disease. Elevated levels of PTHagonist (with respect to PTH antagonist) lead to high bone turnoverdisease (HBT). Elevated levels of PTH antagonist (with respect to PTHagonist) lead to adynamic low bone turnover disease (ALBT). In general,the more medically serious of these two disorders is ALBT due to thehigher risk of soft tissue calcification that accompanies this disorder.In ALBT ectopic tissue calcification results in vascular stenosis(including occlusion of coronary arteries) and aortic rigidity.Therefore, ALBT patients are subject to a higher likelihood of seriousmedical complications due to a circulatory system failure, such asmyocardial infarction than those with HBT. One reason for this is thatit has been difficult in the past to find a reliable therapeutictreatment for ALBT. In addition, the use of therapeutic PTH suppressioncan lead to therapeutic PTH over-suppression which, in turn, leads toALBT. In other words, due to the lack of a reliable indicator, PTHagonist suppressant therapy can inadvertently lead to ALBT due to PTHover-suppression.

Difficulty with implementing PTH suppression therapy can be seen in thesetting of at least ten different PTH target recommendations within thepast decade. The net result of this uncertainty in therapeuticindicators is that the incidence of vascular calcification has beenreported to be 88% for ESRD patients. (See Goodman, W. et al., “CoronaryArtery Calcification in Young Adults with End Stage Renal Disease WhoAre Undergoing Dialysis”; NEJM 2000, May 18; 342:20, 1478-1483.)

A novel finding leading to the present invention is that while the PTHagonist level decreases in response to the administration of PTH agonistsuppressants, PTH antagonist levels may not. Moreover, oversuppressionthrough the use of PTH agonist suppressants may increase the PTHantagonist level. For an untreated renal patient with an elevated PTHagonist level, the administration of a PTH suppressant will alsoinitially suppress the PTH antagonist level. However, as the PTHsuppressant dosage increases, the level of PTH antagonist will reach aminimal level and then start to increase before the level of PTH agoniststops decreasing. To avoid inducing ALBT in such patients, theadministration of PTH suppressant should be adjusted so that the PTHantagonist level reaches and stays about the minimal level mentionedabove. Ordinarily skilled artisans know that this level may vary frompatient to patient, but can determine what is best for a particularpatient through monitoring the PTH level response to therapy.

Preferably, PTH agonist levels are measured directly using an assay thatdoes not detect PTH antagonist either in blood, plasma, or serum. Forexample, Scantibodies Laboratory Whole PTH Assay or ScantibodiesLaboratory CAP Assay are appropriate assays.

PTH antagonist levels should be measured using an assay that eitherdirectly detects PTH antagonist (but not PTH agonist) or indirectlythrough a total PTH measurement that measures the sum of the PTH agonistand PTH antagonist. An indirect measurement subtracts the PTH agonistvalue from the total PTH value, deriving the PTH antagonist value. Thus,one should use a total PTH assay that is designed to have essentially100% cross-reactivity with PTH antagonist and PTH agonist. For example,Scantibodies Laboratory Total Intact PTH Assay or ScantibodiesLaboratory Intact PTH Assay are appropriate assays.

The present method should be used when a PTH suppressant type therapy isused. Applicable treatments include calcium administration, vitamin Dand vitamin D analogue administration, and calcimimetic administration.For untreated ESRD patients, the therapeutic should be administered inincreasing increments from an amount nominal for the selected PTHsuppressant. If the patient is already receiving PTH suppressant typetherapy, one can terminate the therapeutic administration for a timesufficient to allow the patient to return to a relatively non-suppressedstate. The time it takes to return to a relatively non-suppressed statemay range from about 3 to 5 minutes to about 20 minutes up to aboutthree to about six months.

The relatively non-suppressed state may either account for parathyroidgland status and/or bone status. For example, without being bound bytheory, a person may return to a relatively non-suppressed state withrespect to the parathyroid gland status in about 3 to about 5 minutes,frequently ranging up to about 20 minutes. This is because the PTHlevels in a patient, with respect to parathyroid gland status, my changewithin minutes of a change in the dose of PTH suppressant, and a returnto a relatively non-suppressed state might be considered to occur withinminutes. Often times, the return to a relatively non-suppressed state,with respect to the parathyroid gland status, may range from about 10minutes up to one or more hours or days. Also, for example, withoutbeing bound by theory, a person may return to a relativelynon-suppressed state with respect to the bone status in about 3 weeks toabout 6 months, frequently ranging between about 3 to about 6 months.This range may be 3 months, about 4 months, about 5 months or about 6months. Often times, the return to a relatively non-suppressed state mayrange from about 3 or about 4 weeks to about 6 weeks. Also, frequentlythe time it takes to return to a relatively non-suppressed state may bemeasured in days or similar increments within the larger range of weeksor months, e.g., between about one to about 6 or about 7 days within aweek span.

As an alternative to discontinuing PTH type suppressant therapy, if apatient is already receiving PTH type suppressant therapy, one canadjust the amount of therapeutic, up or down, until the level of PTHantagonist is minimized. Frequently, one would take the PTH agonistlevel into account while adjusting therapeutic to minimize the level ofPTH antagonist. The administering physician can easily determine theappropriate dosage on a patient-by-patient basis. The skilled artisanappreciates that the change in dosage for PTH suppressant may differfrom that of another. However, the physician can always be guided by thepresent procedure to affect the best compromise for each individualpatient between PTH over-suppression and abnormally elevated PTH.

C. Parathyroid Hormone Antagonists

In general, a PTH antagonist of the present invention comprises acontiguous portion of human PTH having an amino acid sequence set forthin SEQ ID NO: 1 (PTH₁₋₈₄), or a nucleic acid encoding said portion ofhuman PTH, and said PTH antagonist has the following characteristics: a)the N-terminal amino acid residue of said PTH antagonist starts at anyposition spanning position 2 through position 33 of said PTH₁₋₈₄; b) theC-terminal amino acid residue of said PTH antagonist ends at anyposition spanning position 35 through position 84 of said PTH₁₋₈₄; andc) said PTH antagonist has a minimal length of three amino acidresidues. Preferably, the PTH antagonist is in the form of apharmaceutical composition, which pharmaceutical composition comprisesan effective amount of the PTH antagonist and a pharmaceuticallyacceptable carrier or excipient.

The N-terminal amino acid residue of the PTH antagonist can start at anyposition spanning position 2 through position 33 of said PTH₁₋₈₄. Forexample, the N-terminal amino acid residue of the PTH antagonist canstart at position 2 of the PTH₁₋₈₄. The C-terminal amino acid residue ofsaid PTH antagonist can end at any position spanning position 35 throughposition 84 of said PTH₁₋₈₄. For example, the C-terminal amino acidresidue of the PTH antagonist can end at position 84 of the PTH₁₋₈₄.

In a specific embodiment, the PTH antagonist is a protein or a peptide,or a nucleic acid encoding said protein or peptide, selected from thegroup consisting of PTH₂₋₈₄, PTH₃₋₈₄, PTH₄₋₈₄, PTH₅₋₈₄, PTH₆₋₈₄,PTH₇₋₈₄, PTH₈₋₈₄, PTH₉₋₈₄, PTH₁₀₋₈₄, PTH₁₁₋₈₄, PTH₁₂₋₈₄, PTH₁₃₋₈₄,PTH₁₄₋₈₄, PTH₁₅₋₈₄, PTH₁₆₋₈₄, PTH₁₇₋₈₄, PTH₁₈₋₈₄, PTH₁₉₋₈₄, PTH₂₀₋₈₄,PTH₂₁₋₈₄, PTH₂₂₋₈₄, PTH₂₃₋₈₄, PTH₂₄₋₈₄, PTH₂₅₋₈₄, PTH₂₆₋₈₄, PTH₂₇₋₈₄,PTH₂₈₋₈₄, PTH₂₉₋₈₄, PTH₃₀₋₈₄, PTH₃₁₋₈₄, PTH₃₂₋₈₄, and PTH₃₃₋₈₄. Inanother specific embodiment, the PTH antagonist is a protein or apeptide, or a nucleic acid encoding said protein or peptide, selectedfrom the group consisting of PTH₇₋₆₉, PTH₇₋₇₀, PTH₇₋₇₁, PTH₇₋₇₂,PTH₇₋₇₃, PTH₇₋₇₄, PTH₇₋₇₅, PTH₇₋₇₆, PTH₇₋₇₇, PTH₇₋₇₈, PTH₇₋₇₉, PTH₇₋₈₀,PTH₇₋₈₁, PTH7-82, PTH₇₋₈₃ and PTH₇₋₈₄.

The PTH antagonist can have any suitable length provided that itmaintains its antagonizing activity. For example, the PTH antagonist canhave a length of 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17,18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35,36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53,54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71,72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82 or 83 amino acid residues.

D. Parathyroid Hormone Agonists

In general, a PTH agonist of the present invention comprises acontiguous portion of human PTH having an amino acid sequence set forthin SEQ ID NO:1 (PTH₁₋₈₄), and the PTH agonist has the followingcharacteristics: a) the N-terminal amino acid residue of the PTH agoniststarts at position 1 of the PTH₁₋₈₄; and b) the C-terminal amino acidresidue of the PTH agonist ends at any position spanning position 34through position 84 of the PTH₁₋₈₄.

Without being bound by theory, the N-terminal amino acid residue of thePTH agonist generally starts at position 1 of said PTH₁₋₈₄. For example,the N-terminal amino acid residue of the PTH agonist can start atposition 1 of the PTH₁₋₈₄. The C-terminal amino acid residue of said PTHagonist can end at any position spanning position 34 through position 84of said PTH₁₋₈₄. For example, the C-terminal amino acid residue of thePTH agonist can end at position 84 of the PTH₁₋₈₄.

The PTH agonist can have any suitable length provided that it maintainsits agonizing activity. For example, the PTH agonist can have a lengthof 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50,51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68,69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82 or 83 amino acidresidues.

The present invention is further described by the following examples.The examples are provided solely to illustrate the invention byreference to specific embodiments. These exemplifications, whileillustrating certain specific aspects of the invention, do not portraythe limitations or circumscribe the scope of the disclosed invention.

E. Kits

The invention also provides for kits for carrying out the methods of theinvention. Such kits comprise in one or more containers a means fordetermining and monitoring the level of parathyroid hormone (PTH)agonist in a renal patient having secondary hyperparathyroidism; in oneor more containers, a means for determining and monitoring the level ofPTH antagonist in the patient alone or in combination with other agents;and a means for administering a therapeutic to the patient thatsuppresses PTH agonist whereby the amount of therapeutic administered isadjusted such that PTH agonist levels are decreased and the level of PTHantagonist is minimized. Examples of means for determining andmonitoring PTH agonist levels in a patient comprise a variety of PTHassays further described herein. And, examples of means for determiningand monitoring PTH antagonist levels in a patient comprise a variety ofPTH assays further described herein. Preferred forms may be incombination with sterile saline, dextrose solution, or bufferedsolution, or other pharmaceutically acceptable sterile fluid.Alternatively, useful therapeutic compositions may be lyophilized ordessicated; in this instance, the kit optionally further comprises in acontainer a pharmaceutically acceptable solution, preferably sterile, toreconstitute the complex to form a solution for injection purposes.Exemplary pharmaceutically acceptable solutions are saline and dextrosesolution.

In one aspect, a kit of the invention further comprises a needle orsyringe as a means for administering a therapeutic to a patient,preferably packaged in sterile form, for injecting the composition,and/or a packaged alcohol pad. Instructions are optionally included foradministration of composition by a physician or by the patient.

Other features and advantages of the invention will be apparent from thefollowing detailed description, and from the claims.

The present invention is further described by the following examples.The examples are provided solely to illustrate the invention byreference to specific embodiments. These exemplifications, whileillustrating certain specific aspects of the invention, do not portraythe limitations or circumscribe the scope of the disclosed invention.

EXAMPLES Example 1

A clinical trial was held for ninety ESRD patients. Each patient hadbeen receiving vitamin D suppressant therapy in accordance with themanufacturer's guidelines. Each patient was removed from the therapy fora washout period of four weeks, and this was confirmed by a rise in PTHmeasurements after removal of the therapeutic. PTH maxacalcitol (made byChugai Pharmaceutical Corporation of Tokyo, Japan) suppressant therapywas started after the washout at a constant administration of 5.5 μgintravenously every three days. Blood samples were obtained from eachpatient after the washout (week 0), six weeks after therapy restart(week 6), and twelve weeks after therapy restart (week 12). The sampleswere assayed for PTH agonist levels and PTH antagonist levels using thePTH agonist assay and total PTH assay made by Scantibodies Laboratory,Inc. The samples were assayed for bone specific alkaline phosphataseusing a commercially available immunoassay from Hybritech, Inc. of SanDiego, Calif.

Clinical Results

The results of the assays for the ninety patients are shown in FIG. 2and the following table as median values: TABLE Time Time Time Parameter0 Weeks 6 weeks 12 weeks Total PTH 886 531 525 pg/ml (41% decrease) (1%decrease) PTH agonist 609 361 331 pg/ml (41% decrease) (8% decrease) PTHantagonist 276 160 194 pg/ml (42% decrease) (21% increase) AlkalinePhosphatase 396 344 290 U/ml (13% decrease) (16% decrease)

The Table shows how the PTH suppressant lowered both the PTH agonist andthe total PTH values. After six weeks, the PTH agonist and the total PTHvalues have decreased by 41%. However, after twelve weeks, these valueshave decreased a mere 8% (PTH agonist) and 0.8% (total PTH). Theseresults indicate that the PTH suppressant is having difficulty drivingthe PTH agonist levels down any further. Moreover, the levels of totalPTH and PTH agonist are still commonly regarded as being above normallevels (of less than 37 pg/ml for PTH agonist and less than 65 pg/ml fortotal PTH, see, e.g., Nussbaum S R, et al., “Highly sensitive two-siteimmunoradiometric assay ofparathyrin, and its clinical utility inevaluating patients with hypercalcemia,” Clin. Chem. 1987; 33:1364-67)for non-ESRD patients.

The Table provided above indicates the difference in response of PTHantagonist to the PTH suppressant versus the total PTH and PTH agonistresponse. After six weeks, the PTH antagonist level has decreased by42%, consistent with the PTH agonist levels after six weeks. However,after twelve weeks, the PTH antagonist level increases by 21%, incontrast to PTH agonist levels. The continued use of PTH suppressant atthis level will elevate the PTH antagonist levels. Over a period oftime, the increase in PTH antagonist levels will lead to ALBT andsubsequent vascular calcification (as confirmed in the Table by thefurther drop in alkaline phosphatase at twelve weeks). These patientsneed to have the PTH suppressant dosage lowered to maintain the PTHantagonist at a minimal (in this case 160 pg/ml) level. The medicalbenefit realized in reducing the PTH agonist level beyond a particularlevel, even by a small percentage, is far outweighed by the harm causedby the resulting elevation of the PTH antagonist level. This rise in thePTH antagonist level may cause ALBT by inhibiting osteoclast formation,a necessary component in healthy bone modeling involving bone resorptionand bone turnover.

Example 2

To verify that PTH agonist and PTH antagonist concentrations and the PTHagonist/antagonist ratio accurately discriminate between high and lowbone turnover in renal patients, bone biopsy data is obtained from renalpatients having secondary hyperparathyroidism. See Faugere, M-C, et.al., Kidney Int'l. 2001; 60:1460-68. Bone biopsy data will also verifythat calculation and evaluation of both PTH agonist and PTH antagonistlevel data in renal patients provides a more useful therapeutic andprognostic indicator than evaluation of PTH agonist data alone.

Experimental Design

Patients with a total PTH greater than 200 pg/ml (as measured by anIntact PTH assay), will have PTH agonist and PTH antagonist levels andthe PTH agonist/antagonist ratio determined by Scantibodies® CAP PTHassay(PTH agonist), Scantibodies® Whole PTH assay (PTH agonist),Scantibodies® total intact PTH assay (total PTH) and/or Scantibodies®intact PTH assay (total PTH). Those with a total PTH level greater than400 pg/ml, or between 200-400 pg/ml, and a PTH agonist/antagonist ratioless than 1.5 will undergo double tetracycline bone labeling andoutpatient percutaneous needle biopsy of the pelvic crest bone, underlocal anesthesia.

These patients will also have their bone specific alkaline phosphataseand TRAP (tartrate resistant acid phosphatase) measured through knownmeans. In addition, x-ray data will be obtained from each patientthrough the use of mamographic x-ray techniques known in the art.

Patient Population & Selection Criteria

A large cross-section of hemodialysis and peritoneal dialysis patientswill be evaluated in this study. All dialysis patients will be includedin the initial patient population selection. However, exclusion criteriaincludes patients that are generally considered too ill to participatein the study, those whose anticoagulation cannot be discontinued forabout 48 to 72 hours, and those that are unable to cooperate with studyrequirements. At least 30 to 50 percent of the dialysis population islikely to have PTH values in the range required to be considered forbone biopsy.

Results

Bone biopsy results are correlated with results of the PTH level assaysto verify the accuracy of the PTH agonist/antagonist ratio, versus thePTH agonist measurement alone, in determining a patient's bone turnoverrate. Biochemical and radiological data are used to verify theseresults. As known clinical co-morbidities and therapies have an effecton bone turnover, the effects of these are taken into account inevaluating the data.

The bone biopsy data confirm the efficacy of the PTH agonist/antagonistlevel comparison and therapy is guided based on a comparison of PTHagonist levels with PTH antagonist levels. Therapy is provided to reducePTH agonist levels and to minimize PTH antagonist levels in thepatients.

After twelve months, biochemical and radiological parameters arereanalyzed, and repeat bone biopsies are performed in order to assessthe clinical benefit of the PTH agonist/antagonist ratio analysisapproach.

The above examples are included for illustrative purposes only and arenot intended to limit the scope of the invention. Many variations tothose described above are possible. Since modifications and variationsto the examples described above will be apparent to those of skill inthis art, it is intended that this invention be limited only by thescope of the appended claims.

Citation of the above publications or documents is not intended as anadmission that any of the foregoing is pertinent prior art, nor does itconstitute any admission as to the contents or date of thesepublications or documents.

1-42. (canceled)
 43. A kit for monitoring and guiding therapeuticsuppression of parathyroid hormone in a renal patient having secondaryhyperparathyroidism, the kit comprising: a) means for monitoring thelevel of a parathyroid hormone (PTH) agonist in the patient; b) meansfor monitoring the level of a PTH antagonist in the patient; and c)means for administering to the patient a therapeutic that suppresses PTHagonist, whereby the amount of therapeutic administered is adjusted suchthat PTH agonist levels are decreased and the level of PTH antagonist isminimized.
 44. The kit of claim 43, wherein the therapeutic is selectedfrom the group consisting of vitamin D or vitamin D analogue treatment,calcium treatment, or calcimimetic administration.
 45. The kit of claim44, wherein the Vitamin D analogue comprises paricalcitrol, calcitriol,maxacalcitol, alfacalcidol, calcifediol, or ergocalciferol.
 46. The kitof claim 43, further comprising instructions for use.
 47. The kit ofclaim 43, wherein the PTH agonist comprises a contiguous segment ofhuman PTH having the amino acid sequence SEQ ID NO: 1, wherein saidsegment starts at position 1 of human PTH, and ends at any position fromposition 34 through position 84 of human PTH.
 48. The kit of claim 43,wherein the PTH antagonist comprises a contiguous portion of human PTHhaving the amino acid sequence SEQ ID NO: 1, wherein said portion startsat any position from position 2 through position 33 of human PTH, endsat any position from position 35 through position 84 of human PTH, andhas a minimal length of three amino acid residues.
 49. The kit of claim43, wherein the PTH agonist is a peptide having an amino acid sequenceof human PTH₁₋₈₄ set forth in SEQ ID NO:
 1. 50. The kit of claim 43,wherein the PTH antagonist is a peptide having an amino acid sequence ofhuman PTH7-84 (SEQ ID NO: 7).
 51. The kit of claim 43, wherein the meansfor measuring the PTH antagonist level in a patient comprises means formeasuring a total PTH level in a patient, whereby the PTH antagonistlevel in the patient is determined by subtracting the PTH agonist levelin the patient from the total PTH level in the patient.
 52. The kit ofclaim 43, wherein the means for determining a PTH antagonist levelcomprises an antibody that distinguishes PTH antagonist from PTHagonist.
 53. The kit of claim 43, wherein the means for determining aPTH agonist level comprises an antibody that distinguishes PTH agonistfrom PTH antagonist.
 54. A kit for monitoring and guiding therapeuticsuppression of parathyroid hormone in a renal patient having secondaryhyperparathyroidism, the kit comprising: means for monitoring the levelof a PTH antagonist in the patient; and means for administering to thepatient a therapeutic that suppresses PTH agonist, whereby the amount oftherapeutic administered is adjusted such that the level of PTHantagonist in the patient is minimized.
 55. The kit of claim 54, whereinthe PTH antagonist is a peptide having an amino acid sequence of humanPTH7-84 (SEQ ID NO: 7).
 56. The kit of claim 54, wherein the means fordetermining a PTH antagonist level comprises an antibody thatdistinguishes PTH agonist from PTH antagonist.
 57. The kit of claim 54,wherein the PTH agonist is a peptide having an amino acid sequence ofhuman PTH₁₋₈₄ set forth in SEQ ID NO:
 1. 58. The kit of claim 54,wherein the therapeutic is selected from the group consisting of vitaminD or vitamin D analogue treatment, calcium treatment, or calcimimeticadministration.
 59. The kit of claim 58, wherein the vitamin D analoguecomprises paricalcitrol, calcitriol, maxacalcitol, alfacalcidol,calcifediol, or ergocalciferol.